27. Deutscher Krebskongress

Artikel

Hox- and MMP- gene family members are expressed in freshly frozen and paraffin embedded metastatic colorectal carcinoma – implication for response prediction to a palliative 5-FU containing chemotherapy

Gliederung

Background: An individualized tumour tailored chemotherapy will improve survival of patients with metastatic colorectal carcinoma (CRC). However, this aim requires reliable molecular markers, which will predict tumour response to a specific chemotherapy. Therefore, we analyzed gene expression profiles of liver metastases and primaries of patients with CRC before application of a palliative 5-Fluorouracil (5-FU) based chemotherapy.

Methods: 30 samples of primary CRC and metastatic liver tissue were laser microdissected and the RNA was isolated, amplified and hybridized to Affymetrix HG U-133A microarrays. Additionally, RNA was isolated from formalin-fixed paraffin embedded (FFPE) section of the same tumours and a qRT-PCR was performed for specific gene expression analysis. After surgery of the primary tumour all patients were treated within the framework of a palliative first-line phase III study containing a continuous infusion of 5-FU and folinic acid. Response to chemotherapy was verified by CT scans according to WHO criteria. Gene expression profiles were compared with response to chemotherapy.

Results: By using statistical tests (T-test, Welch, Wilcoxon, Komogorov), support vector machine algorithms and principal component analysis) we identified members of the matrix metalloproteinase (MMP) and HOX gene families as being expressed as well in CRC as well as synchronous liver metastases. A subset of genes consisting of HOXA7, HOXA9, HOXA10, HOXD9, HOXD11 and MMPs -1, -2, -3, -7, -9, and -12 was able to discriminate primary tumours and metastases. RT-PCR experiments approved the results. In a further experiment we extracted RNA from FFPE section of the same CRC and measured the expression of the above mentioned genes by qRT-PCR. We could show, that gene expression results of RNA from FFPE were not only comparable to freshly frozen RNA but were separated (accordingly to the microarray probes) into primary and metastasis by using cluster analysis. Additionally, we compared gene expression profiles generated by qRT-PCR from FFPE tumour probes with clinical response (CR and PR versus SD and PD) to a palliative chemotherapy. It could be shown in this training set, that responders and non-responders could be separated by a "Smallest Informative Biological Signature" (SIBS) containing 4 genes (MMP7, MMP12, HOXA9, HOXD11).

Conclusions: (1) We could show that MMP and HOX gene family members are highly expressed in CRC and synchronous liver metastases; (2) gene expression analysis from freshly frozen material was comparable to qRT-PCR from FFPE material. This will simplify validation of the results in a retrospectively analysis of paraffin embedded tumours; (3) A SIBS of 4 genes was highly effective in predicting response to a palliative 5-FU containing palliative chemotherapy in this training set. Results of the validation from a FFPE test set will be presented on the congress.